Fig. 1

PepO re-programs TAM to the tumoricidal phenotype in vitro. A BMDM or Raw264.7 was polarized into M2 macrophages by IL4(20 ng/ml) and IL-13(20 ng/ml) for 24 h, and then treated with PepO for another 24 h. The transcription level of M1-associated and M2-associated markers were detected by RT-qPCR (n = 3). B BMDM was co-cultured with PY8119 in the presence of PepO or not. The transcription level of macrophage markers was determined by RT-qPCR. Arg-1, CD206, Fizz-1, Ym1, and IL-10 are markers of TAMs, and iNOS, IL-12a, IL-1β are markers of tumoricidal macrophages (n = 3). C The protein level of M1 marker iNOS and M2 marker Arg-1 in TAMs(BMDM + PY8119) and TAMs treated with PepO were evaluated by a western blot analysis. D The expression of pro-inflammatory cytokine TNF-α and IL-6, and anti-inflammatory cytokine IL-10 in TAMs or TAMs treated by PepO were detected using ELISA kit (n = 3). E PY8119 cells were co-cultured with PepO or PBS treated M0 or M2 BMDM, and the percentage of apoptotic PY8119 cells labeled with PI and Annexin V were detected by FACS. F Cell proliferation (Ki-67) was evaluated using Immunofluorescence detection (scale bar = 100 μm). G Heatmap of the DEGs that were uniquely changed in PepO treated M2 macrophages group. H RNA-seq heatmaps showing different gene transcripts (P < 0.05) related to cell killing, phagocytosis and Nitric oxide biosynthetic process in PepO or PBS treated M2 macrophages. Two-way ANOVA with Tukey’s multiple comparisons test was used in (A). One-way ANOVA with Tukey’s multiple comparisons test was used in (B), and (D). Bar graphs represent mean ± SEM, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001